The present invention relates generally to the field of optical spectroscopy and more particularly to the use of optical spectroscopy to observe, analyze and/or characterize biological materials.
The use of optical spectroscopy as a tool for the in vivo and/or in vitro analysis of biological tissues and materials has been studied extensively over approximately the last ten years. This work has demonstrated that optical spectroscopy can be used to provide useful information about both the chemical composition and the morphological structure of biological tissues and materials. Fluorescence spectroscopy and elastic scattering are two of the most common types of optical spectroscopy techniques used to study biological materials. In fluorescence spectroscopy, the photoexcitation of molecules present in a tissue being examined is used to cause the tissue to emit a fluorescence signal that is characteristic of a particular tissue state, e.g., normal vs. benign tumor vs. malignant tumor, etc. In the case of elastic scattering, the absorption and scattering effects that a sample tissue exhibits to illumination provide information that can be used to discern the structure and makeup of the sample. Both of the aforementioned techniques provide a good signal-to-noise ratio for short exposure times to the illuminating light.
An example of fluorescence spectroscopy is disclosed in U.S. Pat. No. 5,131,398, inventors Alfano et al., which issued Jul. 21, 1992, and which is incorporated herein by reference. More specifically, the aforementioned patent describes a method and apparatus for distinguishing cancerous tumors and tissue from benign tumors and tissue or normal tissue using native fluorescence. The tissue to be examined is excited with a beam of monochromatic light at 300 nm. The intensity of the native fluorescence emitted from the tissue is measured at 340 and 440 nm. The ratio of the two intensities is then calculated and used as a basis for determining if the tissue is cancerous as opposed to being benign or normal. The method and apparatus are based on the discovery that, when tissue is excited with monochromatic light at 300 nm, the native fluorescence spectrum over the region from about 320 nm to 600 nm is substantially different for cancerous tissue than for either benign or normal tissue.
Still another type of optical spectroscopy that has been used to study biological materials is Raman spectroscopy. Raman spectroscopy, which arises from inelastic scattering of molecules within a sample, results in the generation of a Raman spectrum, said Raman spectrum containing one or more spectrally narrow peaks. These peaks can be used to identify large biological molecules within the sample or, in some cases, to identify the composition of complex, multicomponent samples. Recently, the use of Raman spectroscopy in the study and diagnosis of diseased tissues has been shown.
For example, in U.S. Pat. No. 5,261,410, inventors Alfano et al., which issued Nov. 16, 1993, and which is incorporated herein by reference, there is disclosed a method for determining if a tissue is a malignant tumor tissue, a benign tumor tissue, or a normal or benign tissue using Raman spectroscopy. Said method is based on the discovery that, when irradiated with a beam of infrared, monochromatic light, malignant tumor tissue, benign tumor tissue, and normal or benign tissue produce distinguishable Raman spectra. For human breast tissue, some salient differences in the respective Raman spectra are the presence of four Raman bands at a Raman shift of about 1078, 1300, 1445 and 1651 cm.sup.-1 for normal or benign tissue, the presence of three Raman bands at a Raman shift of about 1240, 1445 and 1659 cm.sup.-1 for benign tumor tissue, and the presence of two Raman bands at a Raman shift of about 1445 and 1651 cm.sup.-1 for malignant tumor tissue. The aforementioned patent also teaches that, for human breast tissue, the ratio of intensities of the Raman bands at a Raman shift of about 1445 and 1659 cm.sup.-1 is about 1.25 for normal or benign tissue, about 0.93 for benign tumor tissue, and about 0.87 for malignant tumor tissue.
In addition to having applications of the type described above, Raman spectroscopy has also been suggested in recent reports to have the potential to be used in the study and diagnosis of the evolution of precancerous and cancerous lesions in human tissues in vivo.
The Raman scattering process is based on the inelastic scattering of light and typically generates an extremely weak signal, e.g., 10.sup.-6 -10.sup.-14 I.sub.0 where I.sub.0 is the illuminating laser light intensity. The Raman spectrum comprises two spectral components, the Stokes spectrum and the antiStokes spectrum. The Stokes spectrum is located at longer wavelengths than the illuminating laser line whereas the antiStokes spectrum is located at shorter wavelengths than the illuminating laser line. For the above reasons, it can be appreciated that it is essential for Raman spectroscopy that the illuminating laser light be monochromatic and spectrally clean in order for the weak Raman signal emitted by the sample to be observable. At present, Raman spectroscopy is typically performed using a near infrared laser source (e.g., diode laser) and cooled silicon CCD detectors. In general, this type of arrangement yields an acceptable S/N ratio with relatively short integration times.
For Raman spectroscopy to be performed in vivo inside a body, fiberoptics must be used. However, as the illuminating light travels through an optic fiber to a tissue located inside a body, it is common for other spectral components to be generated in the illuminating light due to Raman scattering, fluorescence or like phenomena taking place within the fiber medium. The result of these phenomena is that the light exiting the fiber is not as spectrally clean as necessary. This problem becomes more severe as the length of the fiber increases. As shown in FIG. 1, one way to address this problem is to position a spectrograph, spectrometer or filter array after the fiber. Unfortunately, however, the physical size of these types of filtering arrangements is large (e.g., a few centimeters or larger) and precludes the use of these types of arrangements in many in vivo applications where the fiber is inserted into a working channel of an endoscope, said working channel typically having a diameter about 3 mm.